Determination of analytes in biological fluids in the presence of substances interfering with assays therefor

Abstract

A method is provided, in one embodiment, for the determination of an analyte in a biological fluid sample in the presence of a substance interfering with an assay for the analyte. This embodiment is implemented by using antibodies to cause the selective immunoreaction of at least one of the analyte or the interfering substance and then conducting an assay for the analyte in at least one of the immunoreactants or the non-reactants. Another embodiment provides a disposable reaction device to implement the method. The invention is applicable to the detection of a wide variety of analytes, including cholesterol in a targeted lipoprotein class in the presence of cholesterol in another class; to targeted isozymes of enzymes such as creatine kinase, lactate dehydrogenase, amylase, and alkaline or acid phosphatases in the presence of other isozymes; as well as to targeted immunoglobulins in the presence of non-targeted immunoglobulins.

What is claimed is: 
     
       1. A method for measuring a cholesterol analyte in a targeted lipoprotein class in a biological fluid in the presence of at least one cholesterol-containing interfering substance in another lipoprotein class, comprising: (i) obtaining a disposable reaction device having a reaction chamber, the reaction chamber containing immobilized antibodies which are specific for said interfering substance for removal of the interfering substance from a biological fluid;   (ii) adding the biological fluid sample to the reaction chamber of the device for reaction with said antibodies, wherein the immobilized antibodies bind to the interfering substance without reacting with the cholesterol analyte;   (iii) causing the non-reactants from step (ii), including the cholesterol analyte, to be removed from the reaction device into a collection chamber, wherein the reacted interfering substance remains in the reaction chamber of the reaction device; and   (iv) conducting a clinical assay for the cholesterol analyte on the non-reactants including the cholesterol analyte.   
     
     
       2. A method according to claim 1, wherein a suitable volume of antibodies has been predetermined in relation to a volume of the reaction chamber, the amount of antibodies added to the reaction chamber is sufficient to immuno-react with substantially all of the interfering substance to remove the interfering substance from any anticipated biological fluid sample that may fill the volume of the reaction chamber. 
     
     
       3. A method according to claim 1, wherein step (iii) includes the step of immobilizing the antibodies. 
     
     
       4. A method according to claim 1, wherein step (iii) includes the step of freeze drying the antibodies. 
     
     
       5. A method according to claim 1, wherein the reaction device further includes filter means, for filtering components in the reaction chamber, in fluid communication with the reaction chamber. 
     
     
       6. A method according to claim 5, wherein the reaction chamber includes a first port for receiving a biological fluid sample, a second port for accessing components in the reaction chamber;   wherein the filter means includes a filter disposed in the reaction chamber between the first and second ports;   wherein the device further includes a collection chamber in fluid communication with the second port;   such method further comprising, prior to step (vii), centrifuging the device and allowing the filter to separate the immuno-reactants from the non-reactants which pass into the collection chamber as filtrate.   
     
     
       7. A method according to claim 6, wherein the antibodies are stabilized by immobilizing the antibodies on an insoluble carrier. 
     
     
       8. A method according to claim 6, wherein the antibodies are stabilized by immobilizing the antibodies on a soluble high molecular weight carrier. 
     
     
       9. A method according to claim 6, wherein the antibodies are stabilized by freeze drying. 
     
     
       10. A method according to claim 1, wherein the reaction chamber has first and second ports;   wherein the reaction device includes differential pressure means in fluid communication with the first port for supplying positive and negative pressure differentials to the reaction chamber so that fluid may be alternately drawn into and expelled from the reaction chamber via the second port; and   wherein step (iv) includes drawing the biological fluid sample into the reaction chamber using the differential pressure means.   
     
     
       11. A method according to claim 6, wherein the antibodies are stabilized by immobilizing the antibodies on an insoluble carrier. 
     
     
       12. A method according to claim 10, wherein the antibodies are stabilized by immobilizing the antibodies on a soluble high molecular weight carrier. 
     
     
       13. A method according to claim 6, wherein the step of stabilizing the antibodies are stabilized by freeze drying. 
     
     
       14. A method according to claim 10, wherein the reaction device includes a filter assembly removably attachable to the reaction chamber, the method further comprising prior to step (v), the step of expelling the components in the reaction chamber into the filter assembly using the differential pressure means. 
     
     
       15. A method for measuring a cholesterol analyte in a targeted lipoprotein class in a biological fluid in the presence of at least one cholesterol-containing interfering substrate in another lipoprotein class, comprising: (i) selecting a disposable reaction device having a reaction chamber;   (ii) immobilizing antibodies which are specific for said interfering substance to an interior wall of said reaction chamber;   (iii) adding a biological fluid sample to the reaction chamber of the device for reaction with said antibodies, wherein the antibodies bind to the interfering substance without reacting with the cholesterol analyte;   (iv) separating the non-reactants from step (iii), including the cholesterol, analyte from the interfering substance; and   (v) conducting an assay for the cholesterol analyte on the non-reactants including the cholesterol analyte.   
     
     
       16. A method for measuring a cholesterol analyte in a targeted lipoprotein class in a biological fluid in the presence of at least one cholesterol-containing interfering substance in another lipoprotein class, comprising: (i) obtaining a disposable reaction device having a reaction chamber, the reaction chamber containing antibodies which are specific for said interfering substance for removal of the interfering substance from a biological fluid;   (ii) adding the biological fluid sample to the reaction chamber of the disposable reaction device for reaction with said antibodies wherein said antibodies bind to said interfering substance without reacting with the cholesterol analyte;   (iii) separating non-reactants including the cholesterol analyte from bound interfering substance by filtration; and   (iv) conducting an assay for the cholesterol analyte on the filtrate, wherein the filtrate includes the cholesterol analyte.